Raman spectroscopy for forensic semen identification: Method validation vs. environmental interferences

Proper identification of body fluids at crime scenes is imperative to forensic investigations, as they yield crucial pieces of DNA evidence that lead to subsequent investigation resolution and criminal justice. Nevertheless, body fluid identification can prove a burdensome process, as several ubiquitous substances found at crime scenes often resemble the appearance of a specific body fluid. Therefore, it is extremely important to find a reliable method for body fluid discovery and differentiation among other potential environmental inferences (EIs). In this study, Raman spectroscopy was combined with advanced statistical analysis and used for semen identification. This method has already been proven to be a powerful tool for characterization of body fluid traces for forensic purposes. Twenty-seven EI substances that may (i) yield a false positive (FP) result on routine forensic tests for semen and/or (ii) resemble a semen stain at a crime scene were examined. Raman spectra of EI substances were compared to spectra of main body fluids: semen, blood, saliva, sweat, and vaginal fluid. The random forest algorithm was utilized for differentiating between body fluids and EIs commonly found for semen traces. Using the classification probability threshold of 70%, entire separation of classes was observed. None of the twentyseven EI substances was classified as any body fluid. This confirms a great potential of Raman spectroscopy for nondestructive and rapid identification of semen traces found at a crime scene.